Adenovirus and poxvirus vectors have shown promise for vaccines, but one key hurdle is preexisting immunity to these viruses in humans. The VRC has been investigating the use of DNA priming and alternative routes of immunization to overcome prior immunity. The results demonstrate that priming with a DNA expression vector before immunization with replication-defective adenovirus or poxvirus vectors can overcome the limitations imposed by preexisting immunity to the viral vectors. The VRC is also evaluating the route of delivery of the vaccine (mucosal delivery v. intramuscular) and its effects on the elicitation of cellular and humoral immunity in mice.